The invention concerns an optical system in the beam path of a microscope, specifically a confocal scanning microscope, such that one pinhole diaphragm, the illuminating pinhole, is placed in the illuminating beam between the light source and the object, and another, the detection pinhole, is placed in the detection beam path between the object and the detector.
Optical systems of the type being discussed here have been known in practice for a long time. As examples, we refer only to J. Engelhardt and W. Knebel,"Confocal Laser Scanning Microscopy in Physik in unserer Zeit [Physics in our time], Vol. 24, No. 2, pages 70-78 (1993); J. B. Pawley, in"Handbook of Biological Confocal Microscopy", 2d Edition, pp. 581 ff; and European Patent 0 280 375 B1.
In confocal scanning microscopy in particular, it is necessary to match the diameter of the confocal pinholes to the different magnifications of various microscope objectives. If the pinhole is too large, the resolution along the optical axis is reduced. If the pinhole is too small, on the other hand, the light transmission is reduced; and that is not tolerable, especially in fluorescence applications. Depending on the optical system, the usual pinholes have diameters in the range between 10 .mu.m and 10 mm.
The use of adjustable iris diaphragms in very long beam paths is also already known in practice. These iris diaphragms can be motorized. Their minimum usable diameter is limited to about 0.1 mm, though.
For mechanical-geometrical reasons, and for both mechanical stability and adjustability, compact designs are preferred over long beam paths in order to produce confocal optical systems.
Pinholes with diameters of 5 to 500 .mu.m are used with short light paths. The diameter can be changed by placing several pinholes, each with a fixed diameter, on a disk or wheel. In that way the pinhole diameter can be changed in discrete steps. This is quite difficult because of the small pinhole diameter and the essential positioning accuracy, as such an arrangement is extremely sensitive to adjustment.
A continuously adjustable pinhole is also already known in practice. In this case, two pairs of fine knife edges at right angles can be adjusted, by motorizing, to produce a continuously variable rhomboidal opening between 20 and 500 .mu.m. However, smaller apertures cannot be produced by this known principle because of the micromechanical requirements.